1. Field of the Invention
The present invention relates generally to the field of growth-based detection of microorganisms in sealable containers, such as blood culture bottles. The present invention relates specifically to a system and method for performing optical measurements on a sealable container that can be used to rapidly distinguish positive blood cultures from negative blood cultures, and determine the combination of blood volume and hematocrit in a sealable container.
2. Description of the Related Art
Usually, the presence of biologically active agents such as bacteria or mycobacteria in a patient's body fluid is determined using culture vials. A small quantity of body fluid is injected through the enclosing rubber septum into the sterile vial containing a culture medium. The vial is incubated at 37° C. and monitored for bacterial growth. Known methods preferably detect changes in the carbon dioxide content of the culture bottles, which is a metabolic by-product of the bacterial growth. Changes in the carbon dioxide concentration are usually monitored using chemical sensors disposed on the inner walls of the culture bottles. The chemical sensors respond to changes in the carbon dioxide concentration by changing their color or by changing their fluorescence intensity (see, for example, Thorpe et al. “BacT/Alert: An automated calorimetric microbial detection system”, J. Clin. Microbiol., July 1990, pp. 1608-12; and U.S. Pat. Nos. 4,945,060, 5,217,875, 5,266,486, 5,372,936, and 5,580,784, the entire contents of which are incorporated herein by reference). It has also been suggested to employ carbon dioxide-dependent and/or oxygen-dependent changes in the fluorescence lifetime of fluorescent sensor materials (see, for example, U.S. Pat. Nos. 5,593,854, 5,686,300, 6,074,870, and 6,080,574, the entire contents of which are incorporated herein by reference).
However, these and other known methods generally do not take into account certain factors relevant to the reliability of the detection. These factors include a sufficiently large blood volume for detection and the question of any time delay between extraction and detection.
With regards to the blood volume factor, for the timely and efficient recovery of bacteria from blood samples, it has been found that a sufficiently large blood volume is required (see, for example, Jonsson et al. “Theoretical aspects of detection of bacteremia as a function of the volume of blood cultured”, APMIS 1993 (101:595-601); Mermel et al. “Detection of bacteremia in adults—Consequences of culturing an inadequate volume of blood”, Annals Internal Med 1993 (119:270-272); Arpi et al. “Importance of blood volume cultured in the detection of bacteremia”, Eur J Clin Microbiol Infect Dis 1989 (8:838-842); Isaacman et al. “Effect of number of blood cultures and volume of blood on detection of bacteremia in children”, J Pediatr 1996 (128:190-195); Li et al. “Effects of volume and periodicity on blood cultures”, J Clin Microbiol 1994 (32:2829-2831); Wilson et al. “Controlled evaluation of Bact/Alert standard anaerobic and FAN anaerobic blood culture bottles for the detection of bacteremia and fungemia”, J Clin Microbiol 1995 (33:2265-2270); Wormser et al. “Improving the yield of blood cultures for patients with early Lyme Disease”, J Clin Microbiol 1998 (36:296-298); and Shafazand et al. “Blood cultures in the critical care unit—Improving utilization and yield”, Chest 2002 (122:1727-1736), the entire contents of which are incorporated herein by reference).
The need for employing a large volume of blood in culture bottles arises because, depending on the microorganism species, the number of cell forming units per mL of patient blood may be very low. In practice, and depending on the status of a patient, smaller than optimum amounts of blood are frequently used, which has a negative impact on the reliability of such tests.
Up to this point in time, however, no routine method for the blood volume determination has been introduced into the market. Weighing of the sample container after filling and subtracting an average weight has been used in some studies (see Mensa et al. “Yield of blood cultures in relation to the cultured blood volume in BACTEC 6A bottles”, Med Clin (Barcelona) 1997 (108:521-523), the entire contents of which are incorporated herein by reference). Due to the variability in the weight of individual containers, however, this weighing method suffers from substantial errors, in particular if a low blood volume is involved.
As known to those skilled in the art, the metabolism of the blood itself, independent of any bacterial activity within the container, contributes to the observed increase in the growth of the concentration of carbon dioxide and, may, therefore, cause false-positive culture results. The magnitude of this artifact depends on the blood volume, in that the magnitude is larger the higher the blood volume. As a consequence, much effort is directed toward optimization of sophisticated detection algorithms in an attempt to achieve a shorter time-to-detection while avoiding false-positive culture results (see, for example, Marchandin et al. “Detection kinetics for positive blood culture bottles by using the VITAL automated system”, J Clin Microbiol 1995 (33:2098-2101) and Chapin et al. “Comparison of BACTEC 9240 and Difco ESP blood culture systems for detection of organisms from vials whose entry was delayed”, J Clin Microbiol 1996 (34:543-549), the entire contents of which are incorporated herein by reference).
With regards to the time delay factor, blood culture containers, once they have been inoculated with a patient's blood, should preferably be immediately loaded onto an instrumented blood culture detection system. It is well known, however, that a substantial time delay frequently occurs between the time of inoculation and the time of incubation in an instrument. In some countries, delay times up to 48 hours can be expected. This “delayed container entry” phenomenon may cause serious problems if, during the delay time period, the blood culture container is experiencing elevated temperatures that may already support bacterial growth. As a consequence, the container may be already “positive” (i.e., may contain a fully developed population of microorganisms), when the container arrives at the detection instrument. It would not be advisable, of course, to incubate the container, wait for thermal equilibration to 35 degrees Celsius, and then monitor the container for the occurrence of further bacterial growth. Instead, it would be of great advantage if one could rapidly check the incoming bottle for possible “positivity”, and, if it is already positive, re-direct the container to an instrument that performs a microorganism identification so that appropriate antibiotics can be selected for treating the patient.
In addition, if a “delayed” blood culture container contains already a fully developed microorganism population at the time it arrives at an instrumented detection system, the system's sensor will not see a typical transient. Therefore, the system can only rely on absolute detection signal levels. Depending on the performance characteristics of the sensing method, this represents a higher challenge than under typical monitoring conditions, in particular if culture bottles with highly varying blood amounts are expected.
Accordingly, there exists a need for systems and methods capable of rapidly distinguishing positive blood cultures from negative blood cultures and of determining the combination of the blood volume and the hematocrit of a blood sample after extraction from the human body.